Molecular Determinants of the Ca V β-induced Plasma Membrane Targeting of the Ca V 1.2 Channel *

Ca V β subunits modulate cell surface expression and voltage-dependent gating of high voltage-activated (HVA) Ca V 1 and Ca V 2 α1 subunits. High affinity Ca V β binding onto the so-called α interaction domain of the I-II linker of the Ca V α1 subunit is required for Ca V β modulation of HVA channel gating. It has been suggested, however, that Ca V β-mediated plasma membrane targeting could be uncoupled from Ca V β-mediated modulation of channel gating. In addition to Ca V β, Ca V α2δ and calmodulin have been proposed to play important roles in HVA channel targeting. Indeed we show that co-expression of Ca V α2δ caused a 5-fold stimulation of the whole cell currents measured with Ca V 1.2 and Ca V β3. To gauge the synergetic role of auxiliary subunits in the steady-state plasma membrane expression of Ca V 1.2, extracellularly tagged Ca V 1.2 proteins were quantified using fluorescence-activated cell sorting analysis. Co-expression of Ca V 1.2 with either Ca V α2δ, calmodulin wild type, or apocalmodulin (alone or in combination) failed to promote the detection of fluorescently labeled Ca V 1.2 subunits. In contrast, co-expression with Ca V β3 stimulated plasma membrane expression of Ca V 1.2 by a 10-fold factor. Mutations within the α interaction domain of Ca V 1.2 or within the nucleotide kinase domain of Ca V β3 disrupted the Ca V β3-induced plasma membrane targeting of Ca V 1.2. Altogether, these data support a model where high affinity binding of Ca V β to the I-II linker of Ca V α1 largely accounts for Ca V β-induced plasma membrane targeting of Ca V 1.2.